There are numerous commercially important compounds in the carbohydrate pathway of bacterial strains of the family Enterobacteriaceae including among others 2-KLG, a precursor to ascorbic acid; idonic acid; L-gluconic acid; and 2,5-DKG. Biocatalyic processes have been developed for the production of these compounds. In particular, Anderson et al., (1985, Science 230:144-149) disclose a metabolic pathway for Erwinia herbicola that permits the bioconversion of D-glucose to 2-KLG in a single fermentative step. In this bioconversion, there are a variety of intermediates and one step involves the reduction of 2,5-DKG to 2-KLG which is catalyzed by a recombinantly introduced NADPH-dependent 2,5-DKG reductase. In large scale fermentation of Enterobacteriaceae strains containing recombinantly introduced proteins there remain concerns of a regulatory nature associated with the recombinant nature of the organism.
Vladimir Dellic discloses a summary of a study of organisms for conversion of glucose to ketoacids on internet address “svibor@znanost.hr”. This summary discloses the presence in Erwinia citreus ATCC accession number 31623 of a plasmid of 3.8 kb, designated pPZG500. Dellic discloses the introduction of a tetracycline resistance gene into pPZG500 which was used to clone a 2,5-DKG reductase gene.
There remains a need to develop improved Enterobacteriaceae fermentation strains that have desirable phenotypic characteristics relative to growth conditions and which minimize or eliminate the mobilization properties of resident plasmids under fermentation conditions.